Itch is among the most common dermatological complaints, and has various causes. Approximately 17% of the population suffers from atopic dermatitis, and as much as 80% of the population will experience contact dermatitis in their lifetime. Patients suffering from chronic itch experience significantly decreased quality of life. Despite a pressing need, there are no effective treatments for most severe types of chronic itch. One of the reasons that effective therapies are lacking is that we still do not understand which neurons and neural circuits mediate itch and how they are regulated. In this proposal, we describe experiments that will begin to fill this gap in knowledge. Our lab has previously shown mice lacking the transcription factor Bhlhb5 display pathological itch due to the loss of a subset of inhibitory interneurons in lamina II of the dorsal horn (B5-I neurons). In addition, we have discovered B5-I neurons include a molecularly distinct population that expresses neuronal nitric oxide synthase (nNOS). ). [Importantly, I now have evidence that a subset of projection neurons in lamina I receive strong inhibition from nNOS neurons. These findings suggests that the pathological itch observed in Bhlhb5 mutant mice may be due, at least in part, to the loss of inhibitory nNOS neuron input to projection neurons involved in modulating itch. We therefore we hypothesize that nNOS neurons selectively inhibit the subset of projection neurons that mediate itch. Here I propose to investigate how nNOS inhibitor interneurons modulate somatosensory processing using behavioral, anatomical and electrophysiological approaches. In particular, I will selectively inhibit nNOS neurons in vivo using pharmacogenetics; I will define anatomically the subset of which projection neurons are subject to nNOS- mediated inhibition through confocal microscopy; and I will combine optogenetics and patch-clamp electrophysiology in an intact in vitro preparation to determine how nNOS inhibitory interneurons neurons affect the function of lamina I projection neurons. Overall, these studies will provide important insight into a specific microcircuit in the dorsal spinal cord that may be involved in the modulation of itch. These experiments are a fundamental step in beginning to understand somatosensory circuitry and how we can use this information to develop more effective treatments for chronic pruritus.]